Isolation and propagation of the Ap-Variant 1 strain of Anaplasma phagocytophilum in a tick cell line
Isolation and propagation of the Ap-Variant 1 strain of Anaplasma phagocytophilum in a tick cell line
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DOI：
10.1128/jcm.00478-07
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发表时间：
2007-07-01
影响因子：
9.4
通讯作者：
Kurtti, Timothy J.
中科院分区：
文献类型：
--
作者：
Massung, Robert F.;Levin, Michael L.;Kurtti, Timothy J.
在肩背硬蜱蜱源细胞系ISE6中获得了嗜吞噬细胞无原体菌株Ap-Variant 1的第一个组织培养分离株。两株分离物来自山羊血液样本：一株来自感染了罗得岛州肩胛骨蜱的山羊，另一株来自通过连续接触第一只感染山羊的血液而感染的山羊。从明尼苏达州白尾鹿身上直接分离到8株镰形蜱，首次从蜱中分离到无原体。10个分离株的16S rRNA基因序列均与先前描述的Ap-Variant 1相同，但发现ank基因内部存在差异，表明存在自然变异。明尼苏达蜱无原体阳性率为63.9%；PCR检测的36只蜱中有23只呈阳性。从18只pcr阳性蜱中分离出6株蜱源分离物，分离成功率为33.3%。通过小鼠和山羊的实验感染，研究了罗德岛和明尼苏达州的ap -变体I分离株的宿主亲和性。一种来自明尼苏达州蜱的分离物（MN-61-2）被用于接种幼稚动物，该分离物能够感染一只山羊，但不能感染五只小鼠中的每一只，证实明尼苏达州分离物与来自美国东北部的ap变异I具有相同的宿主嗜性。在ISE6细胞中对Ap-Variant 1分离物MN-61-2进行光镜和电镜观察，发现具有嗜吞噬细胞芽胞杆菌特征的细胞质内含物，在膜结合液泡中存在多形性细菌，并有电子密集和电子明亮两种形式。
The first tissue culture isolates of the unique Anaplasma phagocytophilum strain, Ap-Variant 1, were obtained in the Ixodes scapularis tick-derived cell line ISE6. Two isolates were from goat blood samples: one from a goat infected with L scapularis ticks from Rhode Island and a second from a goat infected by serial passage of blood from the first infected goat. Eight isolates were made directly from L scapularis ticks collected from white-tailed deer in Minnesota and represent the first isolations of an Anaplasma species directly from ticks. Each of the 10 isolates had a 16S rRNA gene sequence identical to that previously described for Ap-Variant 1, but differences within the ank gene were found that suggest natural variation. Prevalence of Anaplasma in the Minnesota ticks was 63.9%; 23 of 36 ticks tested by PCR were positive. Six of the tick-derived isolates were obtained from a set of 18 PCR-positive ticks, for a 33.3% isolation success rate. The conservation of host tropism among the Rhode Island and Minnesota isolates of Ap-Variant I was examined by use of experimental infections of mice and a goat. A Minnesota tick-derived isolate (MN-61-2) was used to inoculate naive animals, and this isolate was able to infect a goat but unable to infect each of five mice, confirming that the Minnesota isolates have the same host tropism as Ap-Variant I from the northeastern United States. Light and electron microscopy of the Ap-Variant 1 isolate MN-61-2 in ISE6 cells showed cytoplasmic inclusions characteristic of A. phagocytophilum with pleomorphic bacteria in membrane-bound vacuoles and both electron-dense and electron-lucent forms.